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Marine Hovakimyan, Rudolf Guthoff, Maria Reichard, Andreas Wree, Ingo Nolte, Oliver Stachs; In vivo Confocal Laser-scanning Microscopy To Characterize Wound Repair In Rabbit Corneas After Collagen Cross-linking. Invest. Ophthalmol. Vis. Sci. 2011;52(14):5305.
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In vivo confocal laser-scanning microscopy (CLSM) has obvious advantages as a non-invasive imaging technique that avoids tissue extraction and provides a unique opportunity for dynamic real-time examination of living corneal layers during the wound healing process. This in vivo CLSM study in rabbits was conducted to provide a quantitative and qualitative analysis of corneal tissue alterations and wound repair over 16 weeks following collagen cross-linking.
Six New Zealand White rabbits underwent riboflavin/UVA cross-linking. In vivo CLSM using a Heidelberg Retina Tomograph equipped with a Rostock Cornea Module was performed preoperatively and at 2, 4, 8, 12 and 16 weeks postoperatively.
From 2 weeks onwards the epithelium demonstrated no abnormalities. Evidence of inflammation (small roundish leucocytes and mature dendritic cells with branched-profile Langerhans cells) was visualized at the level of intermediate, basal epithelial cells and Bowman’s membrane. Nerve fibre regeneration was first noted at 12 weeks. Keratocyte activation and hyperreflective extracellular matrix were observed consistently, but by 16 weeks keratocyte activation was diminished, and the extracellular matrix resumed normal reflectivity; only rare activated keratocytes could be observed occasionally in the anterior stroma by that time. Cell density in the posterior stroma and endothelium regained preoperative values by 4 weeks, although anterior stroma repopulation was still incomplete at 16 weeks.
Complete qualitative and quantitative characterization of corneal wound repair was achieved non-invasively by in vivo CLSM over 16 weeks following collagen cross-linking in rabbits. In terms of assessing the ever-increasing range of cross-linking protocols, in vivo CLSM contributes to minimizing the number of experimental animals required.
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